Re: [ccp4bb] phaser: high z score but no sol
Hi Lisa, Why are you so sure there are 4 molecules in the ASU? There may only be 3 and forcing a fourth molecule is causing lots of clashes. In a similar case, I have seen phaser put two molecules right on top of each other when I forced it to search for too many molecules. In your case I would look at the solution (crystal packing) in coot and see whether the clashes are due to some loops (or domains) which may move upon ligand binding, or whether one of the molecules is at a completely wrong position and if you get a good crystal packing (crystal contacts in all directions) with fewer molecules. You can also start refinement with fewer molecules and see if additional density for missing molecule(s) appears. Good luck! Herman From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of LISA Sent: Thursday, April 19, 2012 8:20 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] phaser: high z score but no sol Hi all, I am trying to solve one structure by molecular replacement with phaser in CCP4. This a complex of a multi-domain domains with small ligand. I have structues of this protein in apo state and with other similar ligand. The space group of this crystal is P21. This crystal should have 4 molecules in ASU. I used the full protein as model but did get any sol and LLG is below zero. Then each domain were used as the search models in phaser with rotation and tranlsation. I can the get high z score (20), and LLG is raising. It looks like I get the right sol, but it have more 50 clashes. Why phaser give wrong sol with so high z socre? Can anyone give me some suggestion to solve my strucutes? Thank you. Best Lisa
Re: [ccp4bb] phaser: high z score but no sol
Would the self rotation map make sense with 4 molecules ? Jürgen Sent from my iPad On Apr 20, 2012, at 2:55, herman.schreu...@sanofi.commailto:herman.schreu...@sanofi.com herman.schreu...@sanofi.commailto:herman.schreu...@sanofi.com wrote: Hi Lisa, Why are you so sure there are 4 molecules in the ASU? There may only be 3 and forcing a fourth molecule is causing lots of clashes. In a similar case, I have seen phaser put two molecules right on top of each other when I forced it to search for too many molecules. In your case I would look at the solution (crystal packing) in coot and see whether the clashes are due to some loops (or domains) which may move upon ligand binding, or whether one of the molecules is at a completely wrong position and if you get a good crystal packing (crystal contacts in all directions) with fewer molecules. You can also start refinement with fewer molecules and see if additional density for missing molecule(s) appears. Good luck! Herman From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of LISA Sent: Thursday, April 19, 2012 8:20 AM To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] phaser: high z score but no sol Hi all, I am trying to solve one structure by molecular replacement with phaser in CCP4. This a complex of a multi-domain domains with small ligand. I have structues of this protein in apo state and with other similar ligand. The space group of this crystal is P21. This crystal should have 4 molecules in ASU. I used the full protein as model but did get any sol and LLG is below zero. Then each domain were used as the search models in phaser with rotation and tranlsation. I can the get high z score (20), and LLG is raising. It looks like I get the right sol, but it have more 50 clashes. Why phaser give wrong sol with so high z socre? Can anyone give me some suggestion to solve my strucutes? Thank you. Best Lisa
Re: [ccp4bb] phaser: high z score but no sol
P21.. You sure about this space group? (very high confidences for space group and laue group in pointless?) F On Apr 19, 2012, at 12:20 AM, LISA wrote: Hi all, I am trying to solve one structure by molecular replacement with phaser in CCP4. This a complex of a multi-domain domains with small ligand. I have structues of this protein in apo state and with other similar ligand. The space group of this crystal is P21. This crystal should have 4 molecules in ASU. I used the full protein as model but did get any sol and LLG is below zero. Then each domain were used as the search models in phaser with rotation and tranlsation. I can the get high z score (20), and LLG is raising. It looks like I get the right sol, but it have more 50 clashes. Why phaser give wrong sol with so high z socre? Can anyone give me some suggestion to solve my strucutes? Thank you. Best Lisa
[ccp4bb] phaser: high z score but no sol
Hi all, I am trying to solve one structure by molecular replacement with phaser in CCP4. This a complex of a multi-domain domains with small ligand. I have structues of this protein in apo state and with other similar ligand. The space group of this crystal is P21. This crystal should have 4 molecules in ASU. I used the full protein as model but did get any sol and LLG is below zero. Then each domain were used as the search models in phaser with rotation and tranlsation. I can the get high z score (20), and LLG is raising. It looks like I get the right sol, but it have more 50 clashes. Why phaser give wrong sol with so high z socre? Can anyone give me some suggestion to solve my strucutes? Thank you. Best Lisa
Re: [ccp4bb] phaser: high z score but no sol
Hi, If you have the correct solution, clashes may be due to loops. It may be an idea to clip these off for the molecular replacement calculations (loops might be shorter in the structure-to-be-solved than in the search model, they may have different conformations in the structure-to-be-solved than in the search model, a common property of loops is that they sometimes have different conformations). You didn't provide much information about what was precisely done wrt molecular replacement calculations, I notice... So what I write is just guesswork. If you have little experience with molecular replacement, it may be a good idea to get advice from a colleague who has plenty and can sit next to you and guide you through the process (again, this is guesswork). HTH, Fred. LISA wrote: Hi all, I am trying to solve one structure by molecular replacement with phaser in CCP4. This a complex of a multi-domain domains with small ligand. I have structues of this protein in apo state and with other similar ligand. The space group of this crystal is P21. This crystal should have 4 molecules in ASU. I used the full protein as model but did get any sol and LLG is below zero. Then each domain were used as the search models in phaser with rotation and tranlsation. I can the get high z score (20), and LLG is raising. It looks like I get the right sol, but it have more 50 clashes. Why phaser give wrong sol with so high z socre? Can anyone give me some suggestion to solve my strucutes? Thank you. Best Lisa
Re: [ccp4bb] phaser: high z score but no sol
Hi, My first guess would be that this crystal possesses translational NCS, and that you're using the old (distributed with current old CCP4) version of Phaser that can't handle tNCS. You can tell if this is the case by looking at whether there's a large off-origin peak in the native Patterson map. If so, then you should try Molrep from the current CCP4 or a newer version of Phaser (recent Phenix release, or upcoming CCP4 release). Regards, Randy Read On 19 Apr 2012, at 07:20, LISA wrote: Hi all, I am trying to solve one structure by molecular replacement with phaser in CCP4. This a complex of a multi-domain domains with small ligand. I have structues of this protein in apo state and with other similar ligand. The space group of this crystal is P21. This crystal should have 4 molecules in ASU. I used the full protein as model but did get any sol and LLG is below zero. Then each domain were used as the search models in phaser with rotation and tranlsation. I can the get high z score (20), and LLG is raising. It looks like I get the right sol, but it have more 50 clashes. Why phaser give wrong sol with so high z socre? Can anyone give me some suggestion to solve my strucutes? Thank you. Best Lisa -- Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical Research Tel: + 44 1223 336500 Wellcome Trust/MRC Building Fax: + 44 1223 336827 Hills RoadE-mail: rj...@cam.ac.uk Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk
Re: [ccp4bb] phaser: high z score but no sol
As Randy pointed out, you should check Patterson map for off-origin peaks. There is also a small chance that you actually have P2 - systematic absences may result from tNCS nearly colinear with crystallographic axis. On Thu, 2012-04-19 at 14:20 +0800, LISA wrote: Hi all, I am trying to solve one structure by molecular replacement with phaser in CCP4. This a complex of a multi-domain domains with small ligand. I have structues of this protein in apo state and with other similar ligand. The space group of this crystal is P21. This crystal should have 4 molecules in ASU. I used the full protein as model but did get any sol and LLG is below zero. Then each domain were used as the search models in phaser with rotation and tranlsation. I can the get high z score (20), and LLG is raising. It looks like I get the right sol, but it have more 50 clashes. Why phaser give wrong sol with so high z socre? Can anyone give me some suggestion to solve my strucutes? Thank you. Best Lisa -- I'd jump in myself, if I weren't so good at whistling. Julian, King of Lemurs
